Apolipoprotein (apo) E plays a key role in lipid metabolism and transport in plasma and the central nervous system. One of the three major human apoE isoforms, apoE4, is a major risk factor for Alzheimer's disease by as yet undefined mechanisms. Our long-range objective is to employ a structure-function approach to establish these mechanisms with the goal of identifying potential therapeutic interventions. This proposal focuses on two major areas related to this goal. First, we will extend the resolution of our x-ray diffraction and small-angle x- ray scattering model of apoE4 bound to phospholipids and compare the structures of apoE4 and apoE3 bound to lipid. Secondly, we will continue our studies in which we link the apoE4 structural property of domain interaction to neurodegeneration and functional and cognitive deficits. The brains of human apoE isoform knock in mice and our Arg-61 apoE mouse model in which domain interaction was introduced, contain lower levels of apoE4 and Arg-61 mouse apoE compared to apoE3 and mouse wild-type apoE, respectively. The lower levels were due to decreased secretion by astrocytes, in which the Arg-61 apoE was selectively degraded, inducing an unfolded protein stress response. Based on these studies, we hypothesize that domain interaction contributes to neurodegeneration through two effects on astrocytes. 1) Domain interaction is recognized as an abnormally folded protein, resulting in an endoplasmic reticulum stress response that leads to downstream effects on key biochemical pathways supporting neuronal integrity. 2) Decreased secretion of apoE4 results in lower levels of cholesterol to support synapse formation and neuronal maintenance. This hypothesis represents a novel paradigm in which apoE4 domain interaction contributes to neurodegeneration and functional and cognitive deficits, involving astrocytes in the absence of additional stressors. We believe that these effects are early events in apoE4 carriers and set the stage for more dramatic responses with the addition of brain stressors (e.g. age, ischemia or A2 toxicity). This paradigm links apoE structure with neurodegeneration and functional consequences and suggests that interference with domain interaction is a viable therapeutic approach. These studies hold the potential to determine the mechanisms by which apoE4 is associated with a high risk for Alzheimer's disease and other forms of neurodegeneration and for identifying therapeutic approaches to reduce this risk. PUBLIC HEALTH RELEVANCE: Apolipoprotein E4 (apoE4) a brain protein, is the major genetic risk factor for Alzheimer's disease although the basis for this association is unknown. Based on our apoE4 structure studies, we propose a new mechanism for this association involving astrocytes, which are cells in the brain that support and maintain nerve cells. These studies hold the potential to identify new therapeutic approaches to reverse the effects of apoE4 on Alzheimer's disease.